CN1558769A - Compositions comprising organic extracts of Geum Japonicum thunb var. and the use thereof - Google Patents
Compositions comprising organic extracts of Geum Japonicum thunb var. and the use thereof Download PDFInfo
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- CN1558769A CN1558769A CNA018238173A CN01823817A CN1558769A CN 1558769 A CN1558769 A CN 1558769A CN A018238173 A CNA018238173 A CN A018238173A CN 01823817 A CN01823817 A CN 01823817A CN 1558769 A CN1558769 A CN 1558769A
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/73—Rosaceae (Rose family), e.g. strawberry, chokeberry, blackberry, pear or firethorn
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P17/00—Drugs for dermatological disorders
- A61P17/02—Drugs for dermatological disorders for treating wounds, ulcers, burns, scars, keloids, or the like
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P19/00—Drugs for skeletal disorders
- A61P19/08—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P9/00—Drugs for disorders of the cardiovascular system
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P9/00—Drugs for disorders of the cardiovascular system
- A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
Abstract
The invention provides an organic extract of Geum Japonicum thunb var. (EGJ), and a pharmaceutical composition comprising the same. The organic EGJ has been identified to show potent dual effects on stimulating early growth (less than 48 hours) of new vessels both in ischemic heart muscles and in infracted heart muscles, and on triggering myocardial regeneration in myocardial infarction, and therefore, may be used in the treatment of ischemic diseases in human being or animals including ischemic heart disease and ischemic limbs, and damaged myocardium associated diseases.
Description
Invention field
The present invention relates to the application of big root grass (Geum Japonicum thunb var.) extract (EGJ), particularly use the ischemic diseases of EGJ organic extract treatment humans and animals, comprise ischemic heart desease and ischemic extremity, and the disease relevant with myocardial damage.
Background of invention
Big root grass (Geum Japonicum thunb var.) be a kind of extensive growth in the Jiangsu Province, China, Jiangxi Province, the plant in Guizhou Province and Yunnan Province.It is a genus that comprises 65 rhizome herbaceous plant or undershrub kind that Fructus Adinae belongs to (Geum), leaf is simple or pinniform is shallow splits, and spends neatly, and described genus comprises such as G.Borisii, G.chiloense, G.coccineum, G.macrophyllum, G.montanum, G.reptans, G.rivale, G.triflorum, G.urbanum and G.japonicum etc.Big root grass (Geum japonicum thunb.) is the perennial herb and the flowering plant of a kind of Rosaceae (Rosaceae).Big root grass whole plant flooding extract is used as the diuretic (Perry, L.M.'s, the medicinal plants in East Asia and Southeast Asia, MIT Press, Cambridge, Mass., 343 pages (1980)) in the Chinese medicine.The known water red bayberry spp.ing plant is rich in tannin.Several hydrolyzable tannin from Geum japonicum, have been isolated, salicylic acid element (gemin) A for example, B, C, D, E and F (Dong, H., Chen, S.X., Kini, R.M., and Xu, H.X., J.Nat.Products.61,1356-60 (1998)).Except tannin, isolated several triterpenoids, comprise 2-hydroxyl oleanolic acid (2-hydroxyoleanolicacid), 2-hydroxyl ursolic acid (2-hydroxyursolic acid), 2,19-dihydroxy ursolic acid (2-dihydroxyursolic acid), 2,3,19,23-tetrahydroxy Folium Vaccinii vitis-idaeae-12 alkene-28 acid-28-O-D-pyranglucoside (Xu, H.X., Zeng, F.Q., Wan, M., and sim, K.Y., J.Nat.Products, 59,643-5 (1996)).
Yet also there is not the open information that is used for the treatment of ischemic diseases and myocardial damage about EGJ.Ischemic diseases, for example coronary heart disease remains the Western countries such as the U.S., and the primary cause of the death in developed regions, Asia such as Hong Kong, and the present primary cause of the death (the AmericanHeart Asseciation that also becomes China, 2001 hearts and apoplexy statistics are upgraded Dallas, Texas:AmericanHeart Asseciation, 2000,1-32).At present feasible Therapeutic Method can only relief of symptoms, even and all present progress are unfortunately arranged, in default of the method for functional neovascularity of growing at the commitment of ischemic myocardium, and the myocardial cell of can not regenerating, so the treatment of ischemic diseases and myocardial damage remains (Uchida, Y., the Yanagisawa-Miwa of difficulty, A., Nakamura, F., Yamada, K., Tomaru, T., kimura, K., and Morita, T., AmHeart be (1995) J.130:1182-1188; Lazarous, D.F., Scheinowitz, M., Shou, M., Epstein, S.E. waits the people, Circulation 91:145-152 (1995); Pu, L.Q., Sniderman, A.D., Brassard, R., Lachapelle, K.J., Graham, A.M., Lisbona, R., andsymes, J.F., Circulation 88:208-215 (1993).According to the research of the past for acute and chronic ischemic animal model or clinical trial, by using such as VEGF, aFGF, bFGF, perhaps the growth factor-induced blood vessel of PDGF needs the time (needing 3-9 week) and be restricted, (8-15) (Risau, W., Nature 386:671-674 (1997); Folkman, J., N.Engl.J.Med.333:1757-1763 (1995); Ware.J.A., Angiogenesis and Cardiovascular Disease (J.A.Ware and M.Simons edit), Oxford University Press, New York, Oxford, 30-59 (1999); Ware, J.A., and Simons, M., Nat.Med.3:158-164 (1997); Arras, M., Ito, W.D., Scholz, D., Winkler, B., Schaper, J., and Schaper, W., J.Clin.Invest.101:40-50 (1998); Banai, S., Jaklitsch, M.T., Casscells, W., Shou, M., Shrivastav, S., Correa, R., Epstein, S.E. and Unger, E.F., Circ.Res.69,76-85 (1991); Arras, M., Mollnau, H., Strasser, R., Ito, W.D., Schaper, J., and Schaper, W., Nat.Biotechnol.16:159-162 (1998); Kurz, H., Wilting, J., Sandau, K., and Christ, B., Microvasc.Res.55:92-102 (1998)), stop up generation very fast (several hours) (Unger, the E.F. of the myocardial necrosis that causes by coronary artery, Shou, M., Sheffield, C.D., Hodge, E., Jaye, M. ， ﹠amp; Epstein, S.E., Am.J.Physiol.264:H1567-1574 (1993); Unger, E.F., Banai, S., Shou, M., Jaklitsch, M., Hodge, E., Correa, R., Jaye, M. ， ﹠amp; Epstein, S.E., Cardiovasc.Res.27:785-791 (1993); And Schlaudraff, K., Schumacher, B., von Specht, B.U., Seitelberger, R., Schlosser, V. ， ﹠amp; Fasol, R., Eur.J.Cardiothorac.Surg.7:637-643 (1993)).Therefore, curative early stage blood vessel takes place and myocardium generation can provide the most effectively replacement therapy strategy, if can be successful, this main treatment that will become many ischemic diseasess and myocardial damage be selected.
Up to now, there are not to induce any similar products of early stage blood vessel generation of heart and the feasible new fully cardiac muscle of regenerating on the world market.According to the research of the past for acute and chronic ischemic animal model or clinical trial, some somatomedin are such as VEGF, aFGF, bFGF or PDGF can promote blood vessel to take place on certain level, but it needs the time (several weeks of needs), and because coronary artery stops up the generation very fast (in several hours) of the myocardial necrosis that causes.Therefore induce early stage blood vessel to become and dwindle cardiac infarction kitchen range area and the important goal of saving diseased tissue.And, even in all latest developments of science and research field, not can be used for the regenerated method of myocardial cell and any medicine yet.
In our nearest research, the extract that we identify a kind of Chinese herbal medicine-big root grass demonstrates stimulation neovascularity early growth (＜48 hours) and the regenerated useful effect of myocardial cell in the rabbit acute myocardial infarction model.Several time-of-weeks that the blood vessel of natural generation of comparing takes place and uses the blood vessel of growth factor-induced to take place, the blood vessel in the inductive cardiac muscle of EGJ takes place only need be less than 48 hours time.Therefore as shown in our animal experiment, this specific characteristic that the inductive early stage blood vessel of EGJ takes place is very useful for the New Policy of developing effective treatment ischemic diseases, particularly when the heart infarction takes place for dwindling the focus of infarct area, it is very important saving the myocardial cell of catching an illness.
To treat the notion that the therapeutic blood vessel of ischemic angiopathy takes place be very attractive idea by improving abiogenous revascularization process.It makes us have an opportunity to make and suffer from the ischemic relevant disease, realizes revascularization more completely such as the patient of coronary heart disease or myocardial infarction.For ischemic heart desease, except preventing, by operation or angioplasty, coronary occlusion can only be alleviated at present.There is not effective medicine can promote the growth (blood vessel generation) of early stage neovascularity at present.And after myocardial infarction took place, the cardiac muscle myocardial cell of tissue regeneration promoting again replaced the myocardial cell of loss.And the scar tissue that replaces necrotic myocardium can also cause the damage of cardiac function.Therefore, obviously need a kind of selectable revascularization method to treat ischemic diseases and the replacement of myocardial cell that promote to damage or loss.Curative blood vessel takes place and myocardium generation will be very the most useful selectable therapeutic strategy, and described strategy may become the many ischemic diseasess of treatment, comprises coronary heart disease and ischemic extremity, and the main treatment of the cardiac muscle of damage is selected.Take place if can realize early stage blood vessel, can substitute many current treatment patterns with having more damaging but more effective therapeutic strategy.
Purpose of the present invention is exactly to be implemented in the commitment growth neovascularity cardiac muscle that replacement damages with the regeneration myocardial cell in the infarcted myocardium by use EGJ.Can EGJ be applied directly to the tip part of the blood vessel blockage of ischemic heart or extremity by the local myocardial injection.For bearing the local myocardial injection and confirming do not have vegetation to form or the patient of tumor, the preparation that can also develop oral administration (tablet) or injecting drug use is in patients'blood or muscle.
The present invention relates to the organic extract of big root grass, especially methanolic extract, described extract stimulates the early stage angiogenesis and the cardiac muscle regeneration of ischemic heart and myocardial infarction, therefore can be used for treating ischemic diseases, such as comprising coronary heart disease, the ischemic heart desease of cardiac infarction, ischemic extremity, myocardial damage and tissue repair.
Summary of the invention
An object of the present invention is to provide a kind of Pharmaceutical composition that comprises organic EGJ, be used for the treatment of patient and ischemia, the disease that tissue injury is relevant with tissue repair.Pharmaceutical composition of the present invention comprises the EGJ and the pharmaceutical carrier of effective dose.
Another object of the present invention provides the application that organic EGJ makes Pharmaceutical composition, described compositions is used for the treatment of and comprises human experimenter and ischemia, the disease that tissue injury is relevant with tissue repair, described application comprises described organic EGJ of effective dose and pharmaceutical carrier or other therapeutic agent, mixes such as the skeletal form generation albumen that is used for the treatment of skeletal defect and disease.
Another object of the present invention provides the method for preparing big root grass extract with organic solvent, and described extract is used for the treatment of experimenter and ischemia, tissue injury and tissue repair diseases associated.Described method comprises described plant is broken into fragment; Described fragment is soaked with organic solvent; Filter product; The evaporation filtrate is to dry.
Another object of the present invention provides and is used for the treatment of experimenter and ischemia, the method for tissue injury and tissue repair relevant disease, the described described organic EGJ that uses effective therapeutic dose to the experimenter that comprises.
Summary of the invention
Fig. 1 has shown that EGJ of the present invention promotes the propagation of HVEC under the various dose in culture.
Fig. 2 has shown according to the present invention with the size of the rabbit heart focus of infarct area after the EGJ processing.
Fig. 3 has shown according to the present invention with forming the neovascularity that is full of hemocyte in the rabbit heart focus of infarct after the EGJ processing.
Fig. 4 has shown that according to the present invention the myocardial cell of PCNA-stained positive in the cardiac infarction kitchen range after handling with EGJ is that the cell mass of central area PCNA stained positive of new regenerated myocardial cell, especially focus of infarct is the most obvious.
Detailed Description Of The Invention
Preparing the process of extract the preferred drying of using from big root grass plant with organic solvent Plant. In the case, plant preferably is broken into powder.
The abbreviation that uses among the present invention " EGJ " if do not particularly point out, refers to big root The extract of the organic solvent as described below of grass plant.
The organic solvent that uses in the method for the present invention is liquid state can be selected from fat in the time of 25 ℃ Family's hydrocarbon or aromatic hydrocarbon, ketone (aldehyde), carboxylic acid, ester and ether, they be not substituted or Replaced by one or more substituting group, perhaps its mixture.
The term " aliphatic hydrocarbon " that the present invention uses refers to alkane, alkene, alkynes and alicyclic ring Family's hydrocarbon generally includes 4-20 carbon.
Term " substituent group " is selected from halogen, hydroxyl, alkyl, thiazolinyl, alkynyl, alkoxyl, alkylthio, alkenyloxy, thiazolinyl sulfo-, alkynyloxy group, alkynyl sulfo-.In these substituent groups, preferred low alkyl group, low-grade alkenyl, low-grade alkynyl, lower alkoxy, low alkyl group sulfo-, rudimentary alkenyloxy, low-grade alkenyl sulfo-, low-grade alkynyl oxygen base, low basic alkynyl sulfo-.Replacing radix can from 1 to 3.
Term " alkyl " refers to the saturated aliphatic hydrocarbon that comprises straight chain and branched group.Preferably, alkyl group has 1 to 20 carbon atom (numerical range whenever; For example " 1-20 " that herein uses is meant the group of mentioning, and refers to alkyl here, may contain 1 carbon atom, 2 carbon atoms, and 3 carbon atoms, or the like, up to comprising 20 carbon atoms).
Comprise 1-6 carbon atom in the term that the present invention uses " low alkyl group, low-grade alkenyl, low-grade alkynyl, lower alkoxy, low alkyl group sulfo-, rudimentary alkenyloxy, low-grade alkenyl sulfo-, low-grade alkynyl oxygen base, low basic alkynyl sulfo-".
In substituted aromatic hydrocarbon, preferably by chlorine and low alkyl group such as chlorobenzene, toluene and dimethylbenzene replace.
In solvent of the present invention, preferably contain the alcohol of 2-18 carbon atom, ether, ketone, carboxylic acid and ester thereof.In these solvents, preferred lower alkyl alcohol more preferably contains the alcohol of 1-4 carbon atom, most preferably methanol.
Soaking step of the present invention can carry out under 10-100 ℃ of temperature.This depends on the boiling point of the solvent that uses.Usually this step can be carried out under the temperature between the boiling point of 10 ℃ and this solvent.Preferably, this step can be carried out at normal temperatures.
In the pharmaceutical composition of the present invention, term " effective content " or " effectively therapeutic dose " are conspicuous for those skilled in the art.It depends on experimenter's age, body weight, health status and administering mode.Usually, the percentage by weight of the extract in the compositions is from 0.01% to 99.99%, preferably from 1% to 99%, more preferably from 10% to 90%, and most preferably from 30% to 70%.
" patient " of Shi Yonging or " experimenter " are meant people or animal herein.Usually, be meant animal or people.Animal can be domestic or wild animal, such as cattle, and horse, pig, deer, rabbit, Canis familiaris L. and other suffer from the animal of the disease of mentioning herein.
" active component " that the present invention mentions is meant the EGJ of disclosed organic solvent among the present invention.
" carrier " of Shi Yonging or " pharmaceutical carrier " are meant a kind of component or composition herein, can accept to mean that it is compatible with other composition of disclosed pharmaceutical composition herein, and can not produce excessively the sufferer that compositions is used and poison, described compositions comprises any and all solvents, diluent and other liquid-carrier, disperse and suspension aids, surfactant, isotonic agent, thickening or emulsifying agent, antiseptic, solid binder, lubricant or the like, and be complementary with required specific dosage form.Because the active component of the present composition is fat-soluble, so when compositions formed liquid preparation, active component at first was dissolved in the alcohol of the carrier that comprises following component: a 2-12 carbon atom, such as ethanol, normal propyl alcohol, isopropyl alcohol, n-butyl alcohol, and benzyl alcohol, glycerol, propylene glycol, N,N-dimethylacetamide, dimethyl sulfoxide (DMSO), olive oil, Oleum Gossypii semen, Oleum Arachidis hypogaeae semen, poppy seed oil, Oleum sesami, soybean oil and other vegetable oil then if desired, are mixed with required preparation.Carrier can also comprise and be selected from the solid that is generally used for pharmaceutical preparation technology, such as gelatin, and lactose, magnesium stearate, Pulvis Talci, calcium carbonate, magnesium carbonate, starch (corn starch and potato starch), and other known in the prior art solid.
Pharmaceutical composition can also comprise one or more compositions that is generally used for pharmaceutical preparation technology, such as surfactant, and promoter, flavoring agent, antiseptic, buffer agent, filler, binding agent, disintegrant, lubricant, etc., those skilled in the art can determine according to the preparation of preparation.
The pharmaceutical composition that the present invention uses comprises whole body and topical formulations, and wherein preferred preparation is suitable for sucking, and is oral, rectum, vagina, intracavity, in the organ, the part comprises the oral cavity, Sublingual, demal, ophthalmic, parenteral, comprise subcutaneous, Intradermal, intramuscular, intravenous, intraarticular and transdermal administration.
Compositions can exist with single or multiple-units dosage form or with bulk form, and can be with the method preparation that is widely known by the people in the pharmaceutical technology.All methods comprise active component EGJ and form the step that the carrier of one or more supplementary elements combines.Preparation is usually by with active compound and liquid-carrier, and the solid carrier of segmentation or closely combine with the two homogeneous and prepare then if desired, is made product the dosage form of wanting.
Being suitable for liquid preparations for oral administration can be with discrete units, such as capsule, and cachets, lozenge, perhaps the form of tablet exists, and per unit comprises the active component of scheduled volume; Exist with powdery or particulate form; Perhaps the form with solution in the aqueous solution or suspension exists; Perhaps the form with oil-in-water and water-in-oil emulsion exists.Liquid preparations for oral administration preferably comprises bubble coating as known in the art to prevent compositions and degrade under one's belt and to provide active component to discharge in small intestinal.The compositions that is suitable for oral administration is included in the lozenge that contains active component in the fragrance matrix, and the fragrance matrix is sucrose and Radix Acaciae senegalis or tragacanth normally; The pastille that perhaps in inertial base such as gel and glycerol or sucrose and Radix Acaciae senegalis, comprises active component.
The compositions that is suitable for parenteral comprises aseptic water of active component and nonaqueous phase injection solution, and described prepared product preferably oozes with predetermined acceptance person's blood etc.These prepared products can comprise antioxidant, buffer agent, antibacterial and make compositions and the isoosmotic solute of intended recipient's blood.The sterile suspensions of aqueous solution and non-aqueous solution can comprise suspending agent and thickening agent.Described compositions may reside in the container of unit dose or multiple-units dosage, in ampoule and bottle, and is stored in before use such as in the sterile liquid carrier such as saline or water.Can be from sterilized powder, granule and tablet are made interim injection solution and suspension.
The compositions that is suitable for topical application according to the present invention preferably adopts ointment, cream cream, lotion, emulsifiable paste, gel, spray, the form of aerosol or oil.
The compositions that is suitable for the transdermal administration can adopt the form of discrete fritter, thus and the long time of receiver's epidermis intimate contact.The compositions that is suitable for the transdermal administration can also be sent by ionotherapy, adopts the form of the optional aqueous buffer solution of active component usually.
Active component preferably is mixed with the form of the dosage unit that is easy to administration to be easy to the form of administration and homogeneous dosage.Use " dosage unit form " herein refers to the physics discrete unit to patient's suitable pharmaceutical compositions of treatment.Each dosage should contain as calculated a certain amount of active component producing needed therapeutic effect, and described active component uses separately or combines with selected pharmaceutical carrier.Typically, the compositions active component is with the form administration of dosage unit, and described dosage unit contains the compositions from about 0.1mg to about 500mg weight, preferably from about 1mg to about 100mg.
Active component of the present invention can with every day every kg receiver body weight approximately from 0.001 to about 120mg dosage level administration, preferred every day every kg receiver body weight from about 0.01 to about 30mg, once a day or for several times, to obtain desired therapeutic effect.
By example, the suitable dose of oral administration should be defined as every kg body weight 30mg every day, and the typical doses of intravenous administration should be defined as every kg body weight 1-10mg every day.
Pharmaceutical composition of the present invention is used for the treatment of and ischemia such as ischemic heart desease and limb ischemia, organ damages such as soft tissue and sclerous tissues's damage, scalpel edge and organization healing such as thigh bone (cervical region) fracture or the relevant disease of low level 1/3 fracture of tibia that require neovascularity to generate.
Be the damage of treatment skeletal muscle, skin injury, scalpel edge and other soft tissue injury, the preparation of preferred topical.Yet for the outside administration of treatment such as burn, emulsifiable paste, cream or powder are the preferential dosage forms of selecting.Be the treatment heart disease, suggestion uses solution to carry out local injection, thus the local concentration of medicine can be very high and action effect better and more effective.If yet patient suffers from optimum or malignant tumor simultaneously, recommend not adopt systemic injection or oral administration, and local injection is safer approach.
Utilize EGJ pharmaceutical compositions treatment and ischemia according to the present invention, in the process of the disease that tissue injury is relevant with organization healing, the aforesaid carrier of EGJ and one or more of effective dose is mixed.In described pharmaceutical composition, the percentage by weight of EGJ is generally from 0.01% to 99.99%.Preferably, the percentage by weight of EGJ is from 1-99% in the compositions.
Embodiment
In the process of screening angiogenic agent from Chinese medicine, the methanolic extract of big root grass demonstrates to stimulate in ischemic myocardial and the infarcted myocardium neovascularity early growth (being less than 48 hours) and causes myocardium regenerated double effects in the myocardial infarction.
Embodiment 1
The dry big root grass whole plant of the 1kg that collects from Chinese Guangdong Province is by pulverize.Then powder at room temperature was soaked in methanol (8L) middle 2-3 hour.The product that obtains is filtered, and filtrate vacuum (50 ℃) is evaporated to drying and obtains powder (50g).
Analyze demonstration EGJ by chromatograph and NMR and mainly contain tannin, triterpenoid, comprise 2-α, 19-alpha-dihydroxy--3-oxygen-12-ursen-28-oic acid, ursolic acid, epimolic acid, maslinic acid, euscaphic acid, tormentic acid, 28-β-D-glucoside of tormentic acid etc.
Embodiment 2
The EGJ powder 0.4g that from embodiment 1, obtains
5% DMSO aqueous solution 100ml
The EGJ powder dissolution in 100ml 5%DMSO aqueous solution to obtain working solution.
The EGJ 0.1g of embodiment 1
Corn starch 0.5g
Lactose 1.87g
Magnesium stearate 0.03g
The EGJ of embodiment 1, corn starch, the lactose homogeneous mixes.Add low amounts of water to mixture.The material that filtration obtains is also dry.In mixture, add magnesium stearate and homogeneous mixing.Use comminutor that synthetic is made bead.Each bead weighs 250mg, and contains the active component of 10mg.
Embodiment 4
The EGJ 80mg of embodiment 1
Sodium chloride 8mg
EDTA 1mg
Sodium phosphate buffer (pH6.5) 10mg
Polyglycol ether 10mg
Distilled water is added to 2ml
All components is dissolved in the distilled water, adds distilled water then to 2ml in mixture.Filter the solution of acquisition to make insufflation with sterilizing filter.
Experiment 1
As shown in Figure 1, use the 5%DMSO solution (embodiment 2) of EGJ to stimulate the human vascular endothelial (HVEC) of cultivation and muscle cell (C2C12 propagation) experiment to generate active to detect its specificity angiogenesis and muscle.In Fig. 1, reference number 1,2 has shown the different culture plates under the same culture conditions respectively with 3.Relatively 5%DMSO contrast, EGJ can obviously promote the propagation of HVEC in the culture in the mode of dose dependent.Zoopery according to us proves that EGJ solution demonstrates the effective double effects for the angiogenesis of infarcted myocardium and muscle generation.In chromatograph detects, EGJ is dissolved in the solvent, and application of sample is on the equilibrated Sephadex LH-20 of 10% methanol post, with gradient water-methanol (increasing the amount of methanol) eluting.Eluting obtains 12 fractions, and detects the HVEC of their promotion cultivations and the double activity of C2C12 cell proliferation.Methanolic extract (EGJ) with specificity double activity is further used for experiment in subsequently the body.Use NMR to measure the homogeneity of the chemical compound that contains in the active part.
Experiment 2
The 5%DMSO solution of this EGJ further is applied to the rabbit heart cerebral infarction models with the effect of research for cardiac muscle.Selecting rabbit is that rabbit has the terminal tremulous pulse that is not connected with small artery as the reason of cardiac infarction model, so the acute coronary obstruction can cause permeable membrane infraction fast.Circle round and prop up (LCX) or a left anterior descending branch tremulous pulse (LAD) in the left side of constriction experiment animal respectively.Respectively the distal end of ligation tremulous pulse is injected EGJ (saturated solution of the 5%DMSO of the EGJ of 0.5ml) or contrast solution (5%DMSO) separately immediately after the connection.The 4th day or the 7th day are put to death animal and are taken out heart and be used for fabric study after the ligation.
Fig. 2 shows that animal (a) the focus of infarct area comparison that the EGJ shown in blue arrow handles according to (b) little 3-5 doubly.
Fig. 3 shows, the animal (a﹠amp that the EGJ shown in blue arrow handles; C) observe many new formation blood vessels that are full of blood cell in the focus of infarct section.Compare down contrast (b﹠amp; D) observe blood vessel still less in.
The heart that Fig. 4 (b) demonstrates EGJ processing in the 7th day has the nuclear new regeneration myocardial cell that has the PCNA stained positive in a large number around the focus of infarct edge, and the some of them tissue becomes cell cluster, is shown with a large amount of blood vessels as black arrow simultaneously and exists.By contrast, shown in Fig. 4 (a), have only the new regeneration myocardial cell of PCNA stained positive seldom around the focus of infarct edge, do not have the new regeneration myocardial cell of central cluster in the contrast heart, and mainly be that fibroblast is present in the focus of infarct.It is myocardial cell with the nuclear that proves the PCNA stained positive that section further uses monoclonal myoglobulin heavy chain (MF20) antibody to carry out histochemical stain.Among Fig. 4, (c) be the high power field of (b) Smalt arrow, and in (d), shown in blue arrow, many new regeneration myocardial cell also are present in EGJ and handle the edge that centers on focus of infarct in the heart around the zone.
When using the monoclonal antibody of anti-PCNA to carry out immunohistochemical staining, have only new regenerative cell's nuclear staining to be positive.Use the Cell immunohistochemical staining method of anti-MF20, have only myocardial cell dyeing to be positive.These dual Cell immunohistochemical staining method and morphological analysis are combined, and new regeneration myocardial cell can be differentiated very reliably.Nuclear PCNA dyeing around the complete not injury of myocardium cell of focus of infarct is negative, and cytoplasmic MF20 dyeing be positive (the brown cell that dyes).
Although the present invention describes and illustration with certain preferred embodiment, other embodiment also is conspicuous to those skilled in the art.Therefore, the invention is not restricted to describe and the particular of illustration, only otherwise breaking away from spirit of the present invention can revise and change it, accessory claim defines four corner of the present invention.
Claims (23)
1. one kind is used for the treatment of experimenter and ischemia, the pharmaceutical composition of the disease that tissue injury is relevant with organization healing, and described compositions comprises the big root grass extractive with organic solvent (EGJ) of effective dose, and a kind of pharmaceutical carrier.
2. according to the pharmaceutical composition of claim 1, wherein said compositions comprises the described extract of from 0.01% to 99.99% weight.
3. according to the pharmaceutical composition of claim 1, wherein said experimenter is human.
4. according to the pharmaceutical composition of claim 1, wherein said organic solvent is selected from lower alkyl alcohol.
5. according to the pharmaceutical composition of claim 4, wherein said low alkyl group comprises 1-6 carbon atom.
6. according to the pharmaceutical composition of claim 5, wherein said organic solvent is a methanol.
7. according to each pharmaceutical composition of claim 1-6, wherein said disease comprises and ischemic heart desease, ischemic extremity, soft tissue or sclerous tissues's damage, scalpel edge, thigh bone (cervical region) fracture or the relevant disease of low level 1/3 fracture of tibia.
8. prepare the method for extract by organic solvent from big root grass plant, described method comprises following steps:
I) described plant is broken into fragment;
Ii) soak described fragment with organic solvent,
The iii) product that ii) obtains of filtration step; With
Iv) filtrate is evaporated to drying.
9. method according to Claim 8, wherein said plant is exsiccant before fragmentation.
10. wherein said step I i according to Claim 8 or 9 method) under the temperature between 10 to 80 ℃, carry out.
11. method according to Claim 8, wherein said step I i) carry out at normal temperatures.
12.EGJ be used for the treatment of and ischemia in preparation, the application in the pharmaceutical composition of tissue injury and organization healing relevant disease, described organization healing need the early stage revascularization of reconstruct blood supply.
13. according to the application of claim 12, wherein said disease comprises coronary heart disease and myocardial infarction.
14. according to the application of claim 12, wherein said disease is the disease relevant with myocardial damage.
15. according to the application of claim 12, wherein said disease is and myocardial cell injury or the relevant disease of forfeiture.
16. according to the application of claim 12, wherein said disease is and the relevant disease of organization healing that comprises the healing of soft tissue and sclerous tissues that described organization healing needs the reconstruct blood supply.
17. according to the application of claim 12, wherein said disease comprises the disease relevant with the ischemic extremity.
18. treatment experimenter and ischemia, the method for tissue injury and organization healing relevant disease, described method comprise the EGJ to described experimenter's administering therapeutic effective dose.
19. according to the method for claim 18, wherein said EGJ carries out administration with the unit dosage form that every day, every kg experimenter's body weight comprised about 0.001 to about 120mg described extract.
20. according to the method for claim 18 or 19, wherein said unit dose comprises a kind of pharmaceutical carrier.
21. according to the method for claim 18 or 19, wherein said disease comprises coronary heart disease, myocardial infarction and ischemic extremity.
22. according to the method for claim 18 or 19, wherein said disease is and the relevant disease of organization healing that comprises the healing of soft tissue and sclerous tissues.
23. according to the method for claim 18 or 19, wherein said disease comprises and myocardial cell injury or the relevant disease of forfeiture.
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| PCT/CN2001/001577 WO2003043645A1 (en) | 2001-11-21 | 2001-11-21 | Compositions comprising organic extracts of geum japonicum thunb var. and the use thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1558769A true CN1558769A (en) | 2004-12-29 |
| CN1258371C CN1258371C (en) | 2006-06-07 |
Family
ID=4574892
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNB018238173A Expired - Lifetime CN1258371C (en) | 2001-11-21 | 2001-11-21 | Compositions comprising organic extracts of Geum Japonicum thunb var. and the use thereof |
Country Status (12)
| Country | Link |
|---|---|
| US (1) | US7572467B2 (en) |
| EP (1) | EP1446134B1 (en) |
| JP (1) | JP4982665B2 (en) |
| CN (1) | CN1258371C (en) |
| AT (1) | ATE481976T1 (en) |
| AU (1) | AU2002221499B2 (en) |
| CA (1) | CA2467895C (en) |
| DE (1) | DE60143132D1 (en) |
| ES (1) | ES2353378T3 (en) |
| HK (1) | HK1068256A1 (en) |
| PT (1) | PT1446134E (en) |
| WO (1) | WO2003043645A1 (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2007048353A1 (en) * | 2005-10-27 | 2007-05-03 | Lead Billion Limited | Pharmaceutical composition and method for neoangiogenesis/revascularization useful in treating ischemic heart diseases |
| WO2010143058A1 (en) * | 2009-06-12 | 2010-12-16 | Generex Pharmaceuticals, Inc. | Compositions and methods for the prevention and treatment of heart failure |
Families Citing this family (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2007049088A1 (en) * | 2005-10-27 | 2007-05-03 | Lead Billion Limited | Method of stimulating growth of functional blood vessels and/or regeneration of myocardium in damaged tissues |
| US20120252744A1 (en) * | 2005-10-27 | 2012-10-04 | Lead Billion Limited | Pharmaceutical Composition and Method for Neoangiogenesis/Revascularization Useful in Treating Ischemic Heart Disease |
| WO2007049089A1 (en) * | 2005-10-27 | 2007-05-03 | Lead Billion Limited | Method of stimulating growth of functional blood vessels and/or regeneration of myocardium in damaged tissues |
Family Cites Families (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CA2209222A1 (en) * | 1997-06-27 | 1998-12-27 | Keng-Yeow Sim | Retrovirus protease inhibitors |
| ATE266636T1 (en) * | 1997-09-09 | 2004-05-15 | Bristol Myers Squibb Pharma Co | BENZIMIDAZOLINONE, BENZOXAZOLINONE, BENZOPIPERAZINONE, INDANONE AND THEIR DERIVATIVES AS FACTOR XA INHIBITORS |
| JPH11236334A (en) * | 1997-12-01 | 1999-08-31 | Nissin Food Prod Co Ltd | Cell adhesion inhibitor |
| JPH11199500A (en) * | 1998-01-07 | 1999-07-27 | Nissin Food Prod Co Ltd | Atopic dermatitis therapeutic agent |
-
2001
- 2001-11-21 DE DE60143132T patent/DE60143132D1/en not_active Expired - Lifetime
- 2001-11-21 PT PT01274733T patent/PT1446134E/en unknown
- 2001-11-21 AU AU2002221499A patent/AU2002221499B2/en not_active Expired
- 2001-11-21 CN CNB018238173A patent/CN1258371C/en not_active Expired - Lifetime
- 2001-11-21 ES ES01274733T patent/ES2353378T3/en not_active Expired - Lifetime
- 2001-11-21 US US10/496,333 patent/US7572467B2/en not_active Expired - Lifetime
- 2001-11-21 CA CA2467895A patent/CA2467895C/en not_active Expired - Lifetime
- 2001-11-21 JP JP2003545326A patent/JP4982665B2/en not_active Expired - Fee Related
- 2001-11-21 WO PCT/CN2001/001577 patent/WO2003043645A1/en active IP Right Grant
- 2001-11-21 AT AT01274733T patent/ATE481976T1/en not_active IP Right Cessation
- 2001-11-21 EP EP01274733A patent/EP1446134B1/en not_active Expired - Lifetime
-
2005
- 2005-01-14 HK HK05100372.3A patent/HK1068256A1/en not_active IP Right Cessation
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2007048353A1 (en) * | 2005-10-27 | 2007-05-03 | Lead Billion Limited | Pharmaceutical composition and method for neoangiogenesis/revascularization useful in treating ischemic heart diseases |
| CN101137387B (en) * | 2005-10-27 | 2011-05-25 | 兆领有限公司 | Pharmaceutical composition and method for neoangiogenesis/revascularization useful in treating ischemic heart diseases |
| WO2010143058A1 (en) * | 2009-06-12 | 2010-12-16 | Generex Pharmaceuticals, Inc. | Compositions and methods for the prevention and treatment of heart failure |
| CN102711768A (en) * | 2009-06-12 | 2012-10-03 | 基因雷克斯制药有限公司 | Compositions and methods for the prevention and treatment of heart failure |
| CN102711768B (en) * | 2009-06-12 | 2014-04-09 | 基因雷克斯制药有限公司 | Compositions and methods for prevention and treatment of heart failure |
| US9050277B2 (en) | 2009-06-12 | 2015-06-09 | Generex Pharmaceuticals, Inc. | Combined Geum japonicum and Centella asiatica extracts for the therapeutic treatment of heart failure |
| US9283255B2 (en) | 2009-06-12 | 2016-03-15 | Generex Pharmaceuticals, Inc. | Compositions and methods for the prevention and treatment of red blood cell coagulation |
| US9629884B2 (en) | 2009-06-12 | 2017-04-25 | Generex Pharmaceuticals, Inc. | Compositions and methods for increasing lifespan and health span |
| US9950019B2 (en) | 2009-06-12 | 2018-04-24 | Generex Pharmaceuticals, Inc. | Compositions and methods for the prevention and treatment of brain diseases and conditions |
Also Published As
| Publication number | Publication date |
|---|---|
| ATE481976T1 (en) | 2010-10-15 |
| AU2002221499B2 (en) | 2007-03-15 |
| US20050064048A1 (en) | 2005-03-24 |
| HK1068256A1 (en) | 2005-04-29 |
| DE60143132D1 (en) | 2010-11-04 |
| EP1446134A4 (en) | 2006-09-27 |
| EP1446134B1 (en) | 2010-09-22 |
| WO2003043645A1 (en) | 2003-05-30 |
| CA2467895A1 (en) | 2003-05-30 |
| JP2005514360A (en) | 2005-05-19 |
| CA2467895C (en) | 2011-09-13 |
| CN1258371C (en) | 2006-06-07 |
| JP4982665B2 (en) | 2012-07-25 |
| US7572467B2 (en) | 2009-08-11 |
| EP1446134A1 (en) | 2004-08-18 |
| AU2002221499A1 (en) | 2003-06-10 |
| ES2353378T3 (en) | 2011-03-01 |
| PT1446134E (en) | 2010-12-17 |
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